Clostridium perfringens is an important Gram-positive anaerobic pathogen of humans and animals that is found ubiquitously in soil and the gastrointestinal tract of vertebrates. It causes a number of histotoxic and enterotoxemic diseases. The species produces an array of extracellular toxins, four of which (alpha, beta, epsilon and iota) form the basis for a toxin-typing scheme, which identifies five toxin types (types A, B, C, D or E) (Songer, 1996). In recent years, a novel toxin, NetB, was shown to be produced by the majority of type A isolates recovered from chickens with necrotic enteritis (NE), an important disease in broiler chicken production, and to play a critical role in NE pathogenesis (Keyburn et al., 2008). This advance raises the possibility that type A strains in a number of other poorly understood but clinically and pathologically distinct enteric diseases of different animal species (Songer, 1996) might contain other as-yet-undescribed necrotizing toxin genes (Nowell et al., 2012).
A number of important toxins, including CPB2 (beta2)-toxin, C. perfringens enterotoxin (CPE, in non-food-borne strains), and all the typing toxins except for alpha-toxin (CPA), are encoded on a conserved family of large plasmids related to the pCW3 tetracycline-resistance plasmid. These plasmids share a conserved core region that includes the transfer of clostridial plasmid (tcp) locus required for conjugation (Parreira et al., 2012; Li et al., 2013). The transmissible nature of key C. perfringens toxin and related virulence genes suggests that virulence of different toxin types can change through plasmid acquisition or loss. However, phylogenetic studies of C. perfringens disease strains also suggest a contribution of the chromosomal background to virulence that varies with the source of the strain. For example, clonality has been described for the majority of bovine type E isolates, for porcine type C isolates, and for isolates from chickens with NE (Jost et al., 2006; Hibberd et al., 2011; Xiao et al., 2012; Lepp et al., 2013).
Clostridium perfringens type A-associated diarrhea and enteric disease in dogs is not well characterized, but its association with disease may range in severity from mild and self-limiting to the fatal acute hemorrhagic diarrhea (Marks, 2012). The acute hemorrhagic gastroenteritis form of disease is marked by severe necrotizing inflammation of the intestinal tract, especially of the small intestine, by hemorrhage and in some cases by rapid death (Prescott et al., 1978; Sasaki et al., 1999). The presence of large numbers of C. perfringens adhering to the necrotic intestinal mucosa is a striking and common feature (Prescott et al., 1978; Sasaki et al., 1999; Schlegel et al., 2012; Unterer et al., 2014). Morbidity may be more common than mortality. Because the infection is not well characterized, no gold standard for diagnosis exists (Marks, 2012). In other species, a recognized predisposing factor for severe C. perfringens enteritis is colostrum- or food-associated trypsin-inhibition which prevents pancreatic trypsin proteolysis of secreted toxin (Songer, 1996), but such a possibility of food-associated infection in canine hemorrhagic gastroenteritis is purely speculative. Although not well characterized, acute haemorrhagic gastroenteritis associated with C. perfringens occurs particularly in small breed dogs (Burrows, 1977).
The role of type A C. perfringens in enteric diseases of horses is also not well understood. There is evidence that CPB2 toxin producing C. perfringens plays a role in the fatal progression of colitis in horses (Herholz et al., 1999; Bacciarini et al., 2003). Vilei and others (2005) demonstrated that some out-of-frame cpb2-positive equine disease isolates produced the CPB2 toxin when grown in sub-inhibitory concentrations of gentamicin. They proposed a feasible direct role of these isolates in antibiotic-associated diarrhea in horses, since treatment with aminoglycoside antibiotics allowed translation of the cpb2 mRNA through induction of a ribosomal frame-shift. Anecdotally, there was an association between the isolation of cpb2-positive C. perfringens from cases of equine colitis and the use of gentamicin in hospitalized diarrheic horses, which ended when the antibiotic was stopped (Vilei et al., 2005). The correlation between CPB2 with severe and sometimes fatal colitis in horses is intriguing but the association remains unproven (Waters et al., 2005). An apparent association has also been noted between the presence of the C. perfringens enterotoxin (CPE) and diarrheal illness in adult horses and in foals, including severe enteric disease (Kanoe et al., 1990; Netherwood et al., 1998; Donaldson and Palmer, 1999; Weese et al., 2001).
Type A C. perfringens with cbp2 and (rarely) cpe genes are commonly found in the feces of healthy foals, whereas type C is seldom found in healthy horses (Tillotson et al., 2002). Considerable work has been done on the important role of type C C. perfringens in neonatal enterocolitis (Traub-Dargatz and Jones, 1993; East et al., 1998). The role of type A in fatal enterocolitis is less clear, but necrosis of the small intestine and colon in 1-14-day-old foals caused by type A C. perfringens has been described in Kentucky (Donahue and Williams, 2002). These isolates possessed cpb2 and cpe (Timoney et al., 2005). Other sporadic case reports associate type A C. perfringens with fatal enterocolitis in neonatal foals (Diab et al., 2011; Hazlett et al., 2011).
WO2013/173910 reported isolation of a putative toxin from Type A C. perfringens, which was termed NetE and was considered as a likely contributor to necrotizing enteritis/haemorrhagic gastroenteritis in dogs and foals.